Periodontitis is an infectious disease, but at present very little is known about the acquisition and transmission of the bacteria that are associated with active disease. A. actinomycetemcomitans is often acquired in childhood or adolescence in the absence of clinical symptoms of disease. The source of infection has not been identified. To track the acquisition of infection it is necessary to have a method which can positively differentiate strains within a species of bacteria. Traditional methodologies, such as immunological or biotyping procedures, or restriction fragment analysis of whole genomic DNA have allowed strains to be classified into groups. However, these techniques have not provided precise identification of individual strains. Technology is available which allows the nucleotide sequence of a specific defined genetic region to be determined. The polymerase chain reaction (PCR) method can be used to amplify any defined region of the genome for analysis. We have focused on the spacer region of the ribosomal RNA gene repeat unit because of this region's potential variability. This will be exploited to provide a fingerprint to be used for strain identification. The ribosomal RNA gene repeat is ideally suited for this type of analysis because it is a well defined genetic unit that contains two highly conserved regions which can be used for priming PCR, and these flank a highly variable region suited for strain differentiation. The ribosomal RNA gene spacer region will be amplified by PCR and sequenced for a series of reference and clinical A. actinomycetemcomitans strains to provide a means of precise strain identification. Using this data the level of genetic diversity between strains isolated from individuals from similar and distant geographic locations will be investigated. This will yield information about the relative level of strain differentiation which may be encountered in a clinical application of the methods. The assay will also be used in a small clinical study to track the transmission of this pathogen within 9 families. The assay will be further developed to allow direct application to plaque samples without the necessity to isolate and culture the bacteria, making it practical to perform more extensive clinical studies. The development of this assay will allow future clinical studies that address questions regarding the ease and common routes of transmission of strains between individuals and the pathogenicity of specific bacterial strains. This will contribute to an understanding of the epidemiology and pathogenesis of periodontal disease and may suggest strategies for treatment and prevention.